Structural biology is the study of the 3-D structure of individual molecules and biological nanoassemblies for the purpose of understanding their function and relating that function to cellular processes. Such studies are used both to provide a physical context to biochemistry as well as to study specific interactions, such as drugs interacting with target molecules, how viruses interact with cells or how biomolecules physically function. This proposal seeks support for continued development of EMAN, an extensive image processing suite for electron microscopists. EMAN is a large package with over 1/2 million lines of code, developed over a decade to produce robust, reliable structural results from TEM data. EMAN's primary use is for a technique known as single particle analysis, which merges thousands to hundreds of thousands of 2-D images of individual molecules/assemblies to produce a high resolution 3-D reconstruction. EMAN has an installed user-base of over 1500 users worldwide, and includes a complete OpenGL based 3-D Graphical User Interface (GUI) built on an extensive library of hundreds of image processing algorithms. This proposal seeks continued support for the development of EMAN and for expanding its capabilities to make it useful to a broader range of researchers. A primary focus in the new cycle is on methods for validation of results to insure reproducibility. While the vast majority of CryoEM structures are completely correct, any field can be made more robust. Part of this will involve connecting EMAN's advanced GUI to other specialized TEM software, which insures that multiple approaches all reach the same scientific conclusions when given the same data. EMAN is also beginning to be used as a component in software projects in other disciplines. This proposal will permit us to give assistance to these developers working to expand EMAN's usefulness to new biological communities. Finally, we will provide continuing support to our existing users, by enhancing documentation and extending EMAN's cross-platform support. Combined, these activities will serve to greatly enhance the productivity of this community and provide useful tools to new groups of NIH funded users. These methods are used in diverse areas of biology, and have expanded our understanding of diseases and conditions as diverse as viral infections, neurodegeneration, heart disease and cancer.